Cell cell interactions play a fundamental role in the genesis of most diseases including cardiovascular disease, cancer and metatasis, and infection and inflammation. The disadvantages and limitations of current antithrombotic therapies and the advantages of the present invention are discussed below in the context of cardiovascular disease, however these discussions are also relevant to other disease states.
Cardiovascular Disease: Treatment and Prevention—State of the Art
Cardiovascular disease is a major cause of morbidity and death in Western societies. It is exacerbated by smoking, hyperlipidemia, hypertension and diabetes. Over the last 40 years, our society has taken multiple steps to reduce cardiovascular disease such as promoting a healthier lifestyle, particularly in regard to smoking and diet. Nonetheless, each year, there are >600,000 percutaneous transluminal coronary angioplasty (PTCA) and surgically invasive procedures, e.g. coronary artery bypass grafting (CABG) in N. America alone, performed in cardiovascular disease patients to improve (cardio)vascular blood flow. While these procedures are beneficial to many patients, the benefits are finite and short-lived, and VW stenosis will reoccur. (RITA Trial Participants. 1993; Kirklin J W et al. 1989). For example, restenosis occurs in 25-30% of patients within 6 months of PTCA despite acute heparin treatment, followed by continuous aspirin (ASA) treatment±oral anticoagulants throughout the 6 month post PTCA period. Heparin is given to accelerate thrombin inhibition by antithrombin III (ATIII), thereby preventing fibrinogen cleavage to fibrin and subsequent fibrin clot formation; ASA is given to acetylate platelet cyclooxygenase, thereby inhibiting thromboxane A2 (TxA 2) synthesis which renders platelets less reactive to prothrombotic stimuli; an oral anticoagulant, e.g. coumadin, is given to decrease the level of vitamin K-dependent procoagulants, thereby decreasing the amounts of procoagulant substrates available for thrombus formation. Thus, the current approach to treat cardiovascular disease is to impair platelet function and/or coagulation as a means to prevent (re)occurrence of heart and blood vessel disease. It does not repair the underlying defect, the latter of which if attempted, might return the patient to a normal healthier state.
The only approaches currently proposed to reverse cardiovascular disease, are the use of lipid lowering agents which decrease the risk of atheriosclerotic lesion formation, and gene therapy. The former approach also has provided some benefit, but again, it does not ‘fix’ the underlying problem. Gene therapy may, in fact, address the issue of repairing the underlying defect(s), but gene therapy for cardiovascular disease is still in its infancy, and not without the risk of complex side effects Libby P and Ganz P. 1997).
A treatment process, which not only corrects the underlying cause of the disease problem but also prevents its onset, is therefore needed. The present invention, which relates to the use of 13-HODE in the regulation of blood cell/VW compatibility per se, offers a mom effective approach than do current antithrombotic therapies to both treating and preventing diseases like cardiovascular disease.
Rationale for Regulating VW Biocompatability
In order to better understand the rationale for regulating VW biocompatibility and its benefits over current antithrombotic treatment practices, the rationale behind the current antithrombotic strategies and their obvious limitations is set out below. This, in turn, will highlight some insights which have led to the concept of regulating VW biocompatibility and the novel approach of using 13-HODE to treat and prevent cardiovascular disease of the present invention.
Regulating VW Biocompatibility
1. Vessel Wall Stenosis & Occlusion:
The problem of vascular stenosis and subsequent occlusion is one of the most important of all medical problems and can produce a very wide range of diseases, the best known of which are coronary, cerebral and peripheral arterial blockage. It is, of course, very difficult and perhaps impossible to study the earliest development of such arterial blockages in humans. People who feel healthy are not inclined to submit to invasive procedures, which, in turn, may detect the onset of the disease before it manifests clinical symptoms. However, it is generally accepted that the processes of restenosis after an artery has been cleared or partially cleared of the occlusive material by a procedure such as angioplasty, is likely in many aspects to be similar to the processes involved in the original development of the problem. Vascular restenosis and occlusion after angioplasty or after vessel wall injury is therefore widely used as a model of the whole series of events involved in both primary and secondary arterial occlusion.
Vascular restenosis thought to occur as a result of a combination of intimal smooth muscle cell (SMC) proliferation, SMC synthesis and secretion of extracellular matrix, and VW remodelling. (Schwartz S M et al. 1995; Strauss B H et al. 1994; Chervu A, Moore W S. 1990; Bocan T M A, Guyton J R. 1985). SMC proliferation per se, occurs in response to the mitogenic effects of thrombin generated at the time of VW injury, to platelet-derived growth factor (PDGF) secreted by platelets which adhere at the site of VW injury, and to mitogens secreted by activated endothelial cells (Bocan T M A, Guyton J R. 1985; Chen L B, Buchanan J M. 1975; Ross R. 1993; Bretschneider E et al. 1997; Fischman D L et al. 1994; Grandaliano G et al. 1998; Stouffer G A et al. 1998).
Polymorphonuclear leukocytes (PMNs) and monocytes/macrophages also invade the VW injury site, activating both coagulation and platelets, thereby augmenting the hyperplasia process (Alexander R. W. 1994; Mallory G A et al. 1939; Mehta J L et al. 1998). Moreover, invading monocytes differentiate into macrophages, ingest lipids, calcium and other blood-derived constituents, thereby forming a more complex atheriosclerotic plaque (Chervu A, Moore W S. 1990; Bocan T M A, Guyton J R. 1985). Thus, there is a multiplicity of cell cell interactions, which trigger and sustain intimal hyperplasia and subsequent restenosis (Ross R. 1993; Schwartz R S. 1998; Cicala C, Cirino G. 1997).
All of these events involve the interactions of blood components with the VW, which under ‘healthy conditions’ occur in response to injury and infection—but do not lead to (cardio)vascular disease. However, when these blood component/VW interactions are exaggerated such as with induced SMC proliferation, platelet/fibrin thrombus formation and VW hyperplasia, (cardio)vascular disease is initiated.
2. VW Injury, Repair and Remodelling:
Recent studies debate the relative contributions of intimal VW hyperplasia per se versus VW remodelling after injury, to subsequent VW restenosis in the clinical setting. Lafont, Post, Mintz et al. argue that W remodelling associated with internal elastic lamina dilation or constriction, contributes more to restenosis after PTCA than intimal hyperplasia (Lafont A et al. 1995; Post M J et al. 1994). The results of the Benestent and STRESS studies are said to be consistent with that argument since increasing the coronary artery diameter with a stent, decreases the need for revascularization (Fischman D L et al. 1994; Grandaliano G et al. 1998). Coats et al agree since there is more SMC-derived collagen (and presumeably more SMGs) in non-stenosed VWs than in stenosed VWs (Coats W D et al. 1997; McGee M P et al. 1995.). The opposite might be expected if hyperplasia was the predominate cause for restenosis. Coats et al. suggested that the failure of our current antithrombotic therapy to inhibit restenosis as effectively as expected, is because that therapy focuses predominantly on inhibiting SMC proliferation. These conclusions, however, do not consider the heterogeneity of proliferating SMCs and their capacity to synthesise various matrices (Frid M G et al. 1997), or the fact that SMC collagen synthesis is affected by the presence (or absence) of the endothelium. Specifically, endothelial cells inhibit SMC protein synthesis, particular type III collagen (Myers P R, Tanner M A. 1998). The opposite is not true. It is also known that the extracellular matrix within a hyperplastic intima of a 1st injury VW, is rich in elastin while the extracellular matrix within the hyperplastic intima of a 2nd injury VW, is rich in collagen (Buchanan M R, Brister S J. 1998), (Capron Let al. 1997.). Moreover, the clinical studies cited above were performed with patients who also required a stent due to the complex nature of their lesions. The restenosis rate in those patients is >4×'s the restenosis rate in PTCA patients who do not require a stent (Antoniucci D et al. 1998). The treatment of PTCA patients who require a stent also differs significantly from the treatment of PTCA patients who do not require a stent (Antoniucci D et al. 1998). These differences are likely to affect subsequent outcome, both at the basic and the clinical end point levels. It is more likely that the relative roles of SMC hyperplasia and VW remodelling in restenosis varies depending on the type of injury and the type of the antithrombotic therapy use.
3. Blood Cell/Injured Vessel Wall Interactions:
Normally, the VW is nonthrombogenic and, therefore, biocompatable with the circulating blood. When the VW is injured, it becomes highly thrombogenic. Injured veins and arteries express tissue factor in both their media and intima. This expression increases over time after injury. Tissue factor expression is minimal in uninjured VWs (Channon K M et al. 1997). Tissue factor expression is enhanced further by PMNs and/or monocyte/macrophages, which invade the injury site. This enhancement is dependent on PMN and/or monocyle/macrophage CD18 integrin expression (Channon K M et al. 1997; McGee M P et al. 1995). VW tissue factor expression activates prothrombin, which is widely distributed throughout VW tissue rich in SMCs (McBane R D et al. 1997). Thrombin upregulates endothelial cell PDGF receptor expression, thereby facilitating SMC proliferation (Grandaliano G et al. 1998; DiCorleto P E, Bowen-Pope D F. 1983), and platelet activation. Activated platelets secrete TxA2 (which is vasoconstrictive), PDGF (which is mitogenic for SMCs) and procoagulants, which exacerbate coagulation (Pakala R et al. 1997). Platelet-related factor Xa/Va activity bound to the injured VW also renders it highly thrombogenic. This latter effect persists for >96 hours (Ghigliotti G et al. 1998). Thus, the multiplicity of these events could be addressed through use of an antithrombotic therapy which targets coagulation, platelet function and inflammation, and which also targets VW thrombogenicity per se. To date, this latter approach is virtually non-existent.
4. Anticoagulant Therapy and VW Hyperplasia:
A number of studies demonstrate that heparin can inhibit experimentally-induced SMC proliferation in vitro and in vivo (Castellot J J Jr et al. 1984; Clowes A W, Clowes M M. 1986; Ferrell M et al. 1992; Hanke H et al. 1992). This suggests that heparin should prevent SMC hyperplasia and subsequent restenosis clinically. However, restenosis occurs clinically despite heparin treatment. It is now recognized that thrombin is protected from inhibition by ATIII and the acceleration of that effect by heparin when thrombin binds to fibrin or other constituents on the injured VW surface (Okwusidi J I et al. 1991; Okwusidi J I et al. 1990; Hogg P J, Jackson G M. 1989; Bar-Shavit R et al. 1989). Moreover, the surface-bound thrombin remains active, contributing to systemic hypercoagulation despite anticoagulant therapy (Ghigliotti G et al. 1998; Brister S J et al. 1993; Wells J et al. 1994; Gill J B et al. 1993). Consequently, surface-bound thrombin can activate platelets, SMC proliferation and further coagulation unchecked. There also is evidence that SMCs, which proliferate in response to repeated injury, are less sensitive to the heparin treatment than SMCs, which proliferate in response to a first injury (Capron L et al. 1997; Geary R L et al. 1995).
5. Antiplatelet Therapy and Hyperplasia
There is little evidence that antiplatelet therapy per se reduces SMC hyperplasia. Clearly, ASA is beneficial in reducing the risks of stroke, myocardial infarction and transient ischemic attacks in patients with a variety of cardiovascular diseases (Aspirin Trialists' Collaboration. 1994). However, the overall risk reduction with ASA, is only □ 25% (Aspirin Trialists' Collaboration. 1994). While this risk reduction is statistically significant, the reduction is modest at best.
Also, ASA may benefit only certain subgroup of patients (Buchanan M R, Brister S J. 1995; Grotemeyer K-H et al. 1993; Grotemeyer K H. 1991). This may be due, in part, to the wide variation in platelet responsiveness to assorted stimuli after ASA ingestion (Mueller M R et al. 1997). The effect of ASA is also finite and has little benefit after 2 years (Aspirin Trialists' Collaboration. 1994).
Alternate antiplatelet agents which block the platelet glycoprotein IIb/IIa (GPIIb/IIIa) receptor have been proposed as superior alternates to ASA. The EPILOG study demonstrated that blocking the GPIIb/IIIa receptor with c7E3 decreases acute ischemic complications in patients undergoing PTCA (The EPILOG Investigators. 1997). Similar results were obtained in the PRISM study using Aggrastat, a non-peptide GPIIb/IIIa antagonist (The Platelet Receptor Inhibition in Ischemic Syndrome Management (PRISM) Study Investigators. 1998). It also has been suggested that the short t½ (half-life) of these compounds may circumvent any bleeding side effect as compared to ASA. However, the bleeding issue still remains controversial. More importantly, like with aspirin, there is little clinical evidence to suggest that long-term hyperplasia is inhibited by these compounds.
Given recent studies, it is not surprising that platelet function inhibitors have little effect on preventing hyperplasia and restenosis. Specifically, Sirois et al made animals thrombocytopenic and then injured their arteries. Thrombocytopenia was sustained for short or long periods of time, and then their platelet counts were restored to normal levels. While the onset of VW hyperplasia was delayed in the long-term thrombocytopenic animals, the potential for SMC proliferation was not inhibited at all. Thus, medial SMC PDGR-β receptor expression was upregulated in all animals despite their being or not being thrombocytopenic. As a result, when the platelet count was restored to normal, SMC proliferation and subsequent intimal hyperplasia were initiated (Sirois M G et al. 1997). These data not only emphasise the need to regulate acute platelet/VW interactions to inhibit chronic intimal hyperplasia, but also suggest that platelet inhibition alone for any finite period of time is unlikely to have a lasting effect.
6. Limitations With the Current Antithrombotic Therapies
While all of the studies cited above, both experimental and clinical clearly indicate clinical benefits with the varied approaches to attenuate the different stages in the development of atherosclerosis, none of these approaches prevent disease onset or facilitate disease regression. Moreover, all of the therapeutic approaches mentioned above act indirectly by compromising coagulation, platelet function and/or injured vessel wall repair. As a result, all patients receiving any form of the currently recommended antithrombotic therapies, are rendered hemostatically dysfunctional, and therefore, at a significant hemorrhagic risk. Thus, there is a clear need for a better antithrombotic approach which leads to the prevention and/or reversal of vascular disease, and which achieves these effects without any adverse side effects.
7. 13-HODE VW Biocompatibility and Hyperlasia
The concept of preventing VW hyperplasia by altering VW biocompatibility has not been considered directly, except perhaps, from the perspective of reducing fat and cholesterol intake in an attempt to reduce VW lipid accumulation and fatty streak formation on the VW. Most attempts have focussed more on the isolation and recombinant synthesis and subsequent utilization of VW constituents to alter blood component properties. For example, there is both experimental and clinical data to suggest that endothelial cell-derived nitric oxide, tissue plasminogen activator and prostacyclin are useful in the treatment of patients at risk of acute thromboembolic events (Gershlick A H et al. 1994; Zerkowski H-R et al. 1993; The GUSTO Investigators. 1993). Their effects, like the antiplatelet and anticoagulant therapies, target platelet function, vessel wall calibre and thrombolysis, thereby compromising hemostasis and coagulation. Moreover, it should be noted that all of these are only produced by the VW following injury or activation, and have little effect on regulating the innate biocompatable properties of a healthy, injured or diseased VW per se.
13-HODE is produced in various cells and tissues of the body, particularly by vascular endothelial cells in healthy vessel walls and by dermal epithelial cells. 13-HODE is formed by the action of an enzyme known as 15-lipoxygenase on the dietary essential fatty acid, linoleic acid. The first step is oxidation of the linoleic acid to give 13-hydroperoxyoctadeca-9Z, 11E-dienoic acid (13-HODE). This is then reduced to 13-HODE. 13-HODE is an important signal transduction molecule which is short-lived and whose synthesis is activated by a variety of different stimuli (Buchanan M R et al. 1985; Haas T A et al. 1990; Weber E et al. 1990; Bertomeu M-C et al. 1990; Brister S J et al. 1990; Buchanan M R, Bastida E. 1988; Cho Y, Ziboh V A. 1994; Mari I. 1998; Kang L-T et al. 1999; Pongracz J, Lund J M. 1999; Friedrichs et al. 1999; C Y, Ziboh V A. 1994). Many of the effects of 13-HODE are mediated by inhibition of protein kinases (PK), particularly PKC and mitogen-activated PK (MAP kinase).
13-HODE which is an oily liquid can be incorporated in much the same way as its parent fatty acid, linoleic acid, into a range of complex molecules including phospholipids and triglycerides (Spiteller G. 1998; Fang X et al. 1999). 13-HODE which is not incorporated into complex lipids is rapidly metabolized by hydrogenation and beta-oxidation (Bronstein J C, Bull A W. 1993; Hecht, Spiteller G. 1998).
Almost all of the studies designed to investigate the effects of 13-HODE involve measuring the effects of altering endogenous 13-HODE production or by adding exogenous 13-HODE (in various forms) to cultured cells in vitro. In the past, there have been few studies, which measure the effects of 13-HODE when given orally or parenterally to animals or humans. This limitation has been due, in part, to the difficulties of making large quantities of 13-HODE and its availability to the scientific community. Consequently, the amount of 13-HODE needed for in vivo studies has been extremely expensive. Second, generally it has been believed that 13-HODE is unstable and readily metabolized, like many signal molecules. As such, it has been thought that it would be a waste of time and money to perform studies involving the oral administration of 13-HODE since none of the orally administered material would be expected to reach its target site of action.
However, there are a few studies which suggest that orally administered 13-HODE has biological relevent effects in vivo. Strews patent describes the use of 13-HODE and other related fatty acids to inhibit aromatase enzymes, which convert androgens to estrogens. The purpose of the treatment is to act on any disease, which is induced by estrogen such as breast cancer, and possibly some types of benign prostatic hyperplasia. However, it should be noted that all of the evidence provided in Streber's patent is based on data obtained in vitro. There are no experiments, which demonstrate that that invention actually works in viva. Moreover, Streber does not provide any details regarding the methods of administration or any practical details as to how the materials might be formulated (although it is stated that ‘tablets or capsules or dragees’ may be used). Finally, the daily dose specified ranges from 100 to 1,000 mg, preferably in the 200 to 500 mg ran. (Streber A S. Hydroxy-octadecadienoic acid for the treatment of estrogen-dependent disease. U.S. Pat. No. 5,102,912, April 1992).
The only study known to us which actually describes any experiments whereby a hydroxy derivative of linoleic acid has been administered orally outside of the experiences with the present invention discussed below, is that of Kaminakai et al (Japanese patent, #7-291862, Nov. 7, 1995). However, they only mention 13-HODE in passing. The actual hydroxy derivative of linoleic acid manufactured for patent use in their experiments, is a different fatty acid; namely, 9-hydroxy-10(E)-12(Z) octadecadienoic acid (9-HODE) which is described in the NMR spectrum shown in FIG. 2 of their patent, and which is stated in the text to be the material actually manufactured and studied in their experiments. These experiments involved the use of 9-HODE given orally to influence the action of Sarcoma 180 tumors implanted in the abdominal cavity of mice. They report an inhibitory effect on the rate of tumour growth, but the minimum effective dose required is 55 mg/kg/day.
The only 13-HODE studies which focus specifically on altering VW biocompatibility to prevent thrombogenesis have been reported by Buchanan et al. Their earlier studies demonstrate that healthy VW cells continuously turn over linoleic acid; i.e. at a time when the endothelium is nonthrombogenic or biocompatable with the circulating blood (Buchanan M R et al. 1985). Intracellular linoleic acid is metabolized to 13-HODE via the lipoxygenase pathway (Haas T A et al. 1990).
They also reported that:                i) endogenous VW 13-HODE plays an important role in regulating VW biocompatibility under both healthy and thrombogenic situations. For example, VW cell thrombogenicity varies inversely with VW 13-HODE levels in both animals and humans. Therefore, increasing endogenous levels of 13-HODE in both animals and humans results in a decrease in VW cell thrombogenicity (Weber E et al. 1990; Bertomeu M-C et al. 1990; Brister S J et al. 1990; Buchanan M R, Brister S J. 1994); and a decrease in platelet/VW interactions following injury. (Weber E et al. 1990; Bertomeu M-C et al. 1990; Buchanan M R, Brister S J. 1994); and        ii) 13-HODE down regulates the ability of the vitronectin receptor to recognise its ligands, thereby decreasing its adhesivity for vitronectin, fibronectin and fibrin(ogen) (Buchanan M R et al. 1998).        
The mechanisms underlying these protective effects of 13-HODE are thought to involve inhibition of protein kinases (PK), particularly PKC and mitogen-activated PK (MAP kinase).
It is important to emphasize that in all of these studies, the aim was always to raise the level of endogenous production of 13-HODE by Undulating its endogenous synthesis or breakdown. None of these studies considered manipulating vessel wall 13-HODE levels by the exogenous administration of 13-HODE. This is clearly demonstrated by a series of experiments involving the administration of Persantine (dipyridamole) which is a phosphodiesterase inhibitor and which was thought might regulate endogenous 13-HODE metabolism.
In these experiments, it was demonstrated that an antithrombotic therapy which involved increasing VW 13-HODE levels, decreased SMC hyperplasia. Rabbits were treated with Persantine (1 mg/kg/day for 7 days) before a 1st or a 2nd VW injury, and then 4 weeks later, intimal SMC hyperplasia was measured. Persantine was given on the basis that it inhibits phosphodiesterase, thereby increasing VW cAMP levels (Weber E et al. 1990; Haas T A et al. 1990). Increasing VW cAMP increased VW linoleic-acid turnover and subsequent VW 13-MODE synthesis, which, in turn, was associated with decreased platelet/VW interactions at the time of injury (Weber E et al. 1990). The Persantine treatment inhibited SMC hyperplasia 4 weeks after VW injury. Platelet function in these animals was unchanged.
These studies are consistent with the discovery that decreasing VW thrombogenicity by increasing VW 13-HODE at the time of injury will attenuate long-term intimal hyperplasia and subsequent VW restenosis. Moreover, this approach did not compromise normal hemostasis and coagulation like the currently used clinical approaches do.
13-HODE, Anti-inflammatory Therapy and VW Hyperplasia
Inflammation has been recognized as an integral part of the thrombotic process as early as 1939 (Mallory G A et al. 1939), yet it is not considered in the rationale for our current antithrombotic therapies. However, there is convincing evidence that attenuating certain inflammatory responses provide a significant benefit. For example, monocytes/macrophages and PMNs express the integrin CD11/CD18 (ICAM), and they release cytokines when activated (Peracchia R et al. 1997; Yasukawa H et al. 1997; Turek J J et al. 1998), which, in turn, stimulate i) β3 integrin expression in other cells such as platelets, endothelial cells and SMCs (Blanks J E et al. 1998; Golino P et al. 1997); ii) tissue factor activation (McGee M P et al. 1995); and iii) PDGF expression (Rubin P et al. 1998; Panek R L et al. 1997). Lipid fractions derived from platelets augment these responses by inducing monocyte/macrophage differentiation and growth (Ammon C et al. 1998). Macrophages interacting with the injured vessel wall, accumulate lipid, leading to the formation of a more complex atherosclerotic lesion (Ross R. 1993; Post M J et al. 1994). Blocking monocyte/macrophage ICAM expression reduces VW hyperplasia significantly (Golino P et al. 1997; Nageh M R et 1997; Natori S at al. 1997). Others have found that radiation (90Sr/Y or 192Ir) at doses, which selectively impair monocyte/macrophage function also, decreases VW hyperplasia in both rodent and rabbit models (Rubin P et al. 1998; Panek R L et al. 1997; Williams D O. 1998; Kipshidze N et al. 1998). In a preliminary clinical study, the SCRIPPS trial using endovascular radiation, the restenosis rate in 35 patients undergoing PTCA who also required a stent was 11%, significantly less than the 37% restenosis rate seen in comparable non-irradiated controls (Williams D O. 1998). These data provide direct evidence, which suggests that altering inflammatory responses also affect intimal hyperplasia and subsequent VW restenosis.
Studies by Buchanan et al suggest that the culprit inflammatory cell is not the PMN. In fact, PMNs may attenuate the vessel wall thrombogenicity by providing a source of 13-HODE at the site of blood cell/VW interactions at the time of VW injury (Buchanan M R. 1989; Buchanan M R et al. 1993). Others have argued that PMN-derived oxygen radicals promote ischemia-related damage (Shen J et al. 1996), but this has not been linked to long-term hyperplasia. PMNs also secrete, a nitric oxide-like factor, which inhibits platelet function and vasoconstriction (Cerletti C et al. 1992). Monocytes/macrophages, on the other hand, normally do not synthesize 13-HODE (Shen J et al. 1996; Shen J et al. 1995). Interestingly however, Shen et al upregulated 15-lipoxygenase in differentiated macrophages and found that 13-HODE synthesis increased. This increase was associated with decreased macrophage lipid accumulation. Fan et al. also found that macrophages enriched with linoleic and gamma linolenic acids (substrates for 13-HODE and PGE1, respectively), stimulate intracellular SMC cAMP which, in turn, decreases SMC proliferation (Fan Y Y et al. 1997). Finally, 13-HODE inhibits PAF (platelet activating factor)-induced PMN and monocyte/macrophage degranulation and ICAM expression (Cerletti C et al. 1992), thereby preventing further integrin-dependent cell cell and cell ligand interactions.
Recent Studies with 13-HODE
The above data suggest that endogenous VW 13-HODE plays an important role in regulating VW biocompatibility under both healthy and thrombogenic situations, and that 13-HODE is therefore a useful antithrombotic agent. However, any progress in developing that concept has been thwarted by the lack or absence of any agent which would directly upregulate 13-HODE synthesis by VW cells, PMNs or other relevent cells. While Persantine has been a useful tool to generate preliminary data, it is a weak and reversible inhibitory of phosphodiesterase. Supplementing a diet of cardiovascular diseased patents with linoleic acid (the substrate for 13-HODE)) also has been useful to demonstrate the benefit of elevating VW 13-HODE levels. However, that approach requires the patients to ingest a daily dose of 20 grams or more of linoleic acid-rich capsules, and the treatment is not without its unwanted side effects, including an increased caloric intake.
Surprising Results
To date, researchers working in the field have concentrated on the idea of maintaining healthy endothelial cell function either by regulating the endogenous production of 13-HODE, avoiding factors which suppress its synthesis and/or providing agents containing linoleic acid which may enhance the synthesis of 13-HODE. It was thought that only trivial amounts of purified 13-HODE would reach the target site of the VW endothelium if administered orally, and would, therefore, be biologically inactive. Recently, these ideas were tested and, surprisingly, proven wrong. Specifically, it was found that orally administered 13-HODE does reach its intended targets and is biologically active. In addition, suitable vehicles in which 13-HODE can be administered orally were identified. And most amazingly, the beneficial effects of orally administered 13-HODE are achieved with unexpectedly low doses.
These and other objects and advantages of the invention will be apparent to those skilled in the art from a reading of the following description and appended claims.